Man and other eukaryotic organisms possess internal cellular mechanisms which are manifested as daily (Circadian) rhythms. Biological rhythmicity is important because it involves regulatory processes associated with many fundamental cellular reactions, ranging from cell division and gene expression to enzyme activity. It is also of fundamental importance in human physiology. In both psychiatry and medicine it is known that circadian rhythmicity is involved in the timing of bodily functions, work-rest cycles, jet lag, sleep, drug tolerance, drug efficacy, and even aging and longevity. This research program is concerned with the elucidation of the basic cellular and molecular mechanisms involved in circadian rhythmicity. Recent and current research with an enzyme (luciferase) in the model circadian system of the unicell Gonyaulax has shown that the amount of enzyme actually increases and decreases in concert with the in vivo rhythm. This privides (for the first time) a specific molecular correlate of a circadian rhythm, a measure of a product species controlled by the cellular oscillator. We now aim to determine whether it is transcriptionally or translationally controlled. To accomplish this we will clone luciferase cDNA and use this as a hybridization probe to determine luciferase mRNA levels at different circadian phases. Inhibitors of 80S ribosome protein synthesis cause phase shifts in the circadian rhythm; the effects of these drugs on protein synthesis in Gonyaulax will be compared with effects on the rhythm. The effects of other types of drugs and inhibitors will also be examined for their phase-shifting ability and biochemical effects. Recent results give some evidence for the involvement of cAMP in circadian control; we will therefore study phosphodiesterase inhibitors, protein kinases and phosphorylation patterns in Gonyaulax. Mathematical modeling predicts a metastable arrhythmic "singular" state. We have recently succeeded in generating apparent arrhythmicity by drug treatment; this occurs only at a specific (moderate) dose applied at a specific time. We aim to study the cellular and biochemical status of the rhythmic system in such arrhythmic cells.